Material Basis And Mechanisms For Neuroprotective Effects Of Maca(Lepidium Meyenii Walp.)

Alzheimer’s disease,Parkinson’s disease,and depression are becoming increasingly common in the global population.The drugs presently used to treat neurological diseases often have strong toxic and side effects.Identifying food with neuroprotective effects and safety or natural active ingredients from food has become a research hot spot for the development of drugs or functional food to prevent and treat neurological diseases.Studies at home and abroad have shown that maca(Lepidium meyenii Walp.)has neuroprotective potential,but its active substance basis and mechanisms are unclear.This study aimed to investigate the protective effects of hypocotyl of maca on corticosterone(CORT)-induced neurotoxicity,to determine the main active substance basis by the neuroprotective effects in vivo and in vitro,and to elucidate its action mechanisms.The main results are as follows.(1)Ethanol extract from maca(EEM)could improve CORT-induced neurotoxicity in rats.A neurotoxic model was established by subcutaneous injection of CORT(40 mg/kg·BW)for 21 days.Depressive behaviors(percentage of sucrose consumption,immobility time in forced swimming test,and total distance in open field test)were observed.The levels of brain-derived neurotrophic factor(BDNF),the contents of tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6),and the number of positive cells of doublecortin and bromodeoxyuridine in hippocampus were measured.Behavioral experiments showed that EEM(200 and 400 mg/kg·BW)could remarkably improve the depressive behaviors;enzyme-linked immunosorbent assay(ELISA)showed that EEM could considerably reduce hippocampal neuroinflammation and increase hippocampal neurotrophy;immunofluorescence assay showed that EEM could remarkably increase the number of hippocampal newborn neurons.(2)Macamides may be the neuroprotective material basis of maca.The protective effects of EEM and four different polar fractions were investigated with a CORT-induced PC12 cell injury model.EEM,petroleum ether extract from maca,and ethyl acetate extract from maca could improve PC12 cell viability,and petroleum ether extract from maca was more effective.However,N-butanol extract from maca and aqueous extract from maca could not increase PC12 cell viability.High-performance liquid chromatography(HPLC)revealed that EEM,petroleum ether extract from maca,and ethyl acetate extract from maca contained 954.31,4802.07,and 127.56 mg/100 g total macamides,respectively.N-butanol extract from maca and aqueous extract from maca did not contain macamides.The blood-brain barrier(BBB)permeabilities of 45 compounds in maca were predicted with Discovery Studio software.The lipid/water partition coefficients(Alogp98)of macamides ranged from 3.888 to 11.204,among which N-(3-methoxybenzyl)-9Z,12 Z,15Z-octadecatrienamid(M 18:3)and four other kinds of macamides showed the strongest BBB permeability.The neuroprotective effects of 12 major macamides were evaluated using the CORT-induced PC12 cell-injury model.Unsaturated macamides exerted better protective effects than saturated macamides,and M18:3 had stronger protective activities toward CORT-induced damage.(3)The neuroprotective cellular mechanisms of M 18:3 may involve the reduction of reactive oxygen species(ROS),the inhibition of mitochondrial apoptosis,and the activation of protein kinase B(Akt)and c AMP-response element binding(CREB)protein phosphorylation.The protective effects of M 18:3 were investigated with a CORT-induced PC12 cell-injury model.M 18:3(25 μM)significantly increased the cell viability,decreased the lactate dehydrogenase release rate,decreased the level of intracellular ROS,decreased the ratio of Bax/Bcl-2,and decreased the expression levels of Bax,cytochrome c(Cyt-C),cleaved-caspase-3,and cleaved-poly(ADP-ribose)polymerase(PARP).(4)The neuroprotective effect of M 18:3 was related to the promotion of neurogenesis,the increase in density of hippocampal dendritic spines,and the increase in synapsis-associated proteins.A neurotoxic model was established by subcutaneous injection of CORT(40 mg/kg·BW)for 21 days.The density of hippocampal neurons was calculated.The numbers of positive cells of hippocampal doublecortin,neuronal nuclei protein,and bromodeoxyuridine were measured.The morphological changes of hippocampal neurons(density of dendritic spines,dendritic length,and area and volume of dendritic cell bodies)were observed.The expression levels of synaptophysin,synapsin I,and postsynaptic density protein 95 were measured.Nissl staining showed that M 18:3(5 and 25 mg/kg·BW)could remarkably improve the CORT-induced decrease in hippocampal neuron density;immunofluorescence analysis showed that M 18:3 could considerably promote hippocampal neurogenesis;Golgi staining showed that M 18:3could remarkably improve the CORT-induced changes in hippocampal dendritic structure;Western blot showed that M 18:3 could considerably increase the expression levels of synaptic-structure-related proteins in the hippocampus.In conclusion,ethanol extract from maca can improve CORT-induced neurotoxicity in rats and was thus a safe and natural active extract with neuroprotective effects.Cell and animal experiments showed that macamides can promote nerve cells’ survival and growth,which are the main neuroprotective material basis of maca.The neuroprotective mechanisms of macamides involved the inhibition of neuronal mitochondrial apoptosis,the promotion of hippocampal neurogenesis,and the increase in BDNF and synaptic-protection-related proteins in the hippocampus.EEM and its rich macamides,especially N-(3-methoxybenzyl)-9Z,12 Z,15Z-octadecatrienamid,had good potential to develop functional foods and drugs to prevent and treat neurological diseases.